Journal: Ecotoxicology and environmental safety

Article Title: Additive cardiotoxicity of a bisphenol mixture in zebrafish embryos: The involvement of calcium channel and pump.

doi: 10.1016/j.ecoenv.2023.115225

Figure Lengend Snippet: Fig. 4. Effects of BPs on cardiomyocyte development. Representative confocal microscopy image of TG(myl7:NLS-mCherry) ZFEs at 48 hpf exposed to 0.05% DMSO (A) or Mix 2 (BPA 8.7 µM + BPF 18 µM + BPAF 1.3 µM + BPB 4.2 µM), which induced approximately 40% heart rate reduction (B). BP exposure did not affect the cardiomyocyte count (C), based on the t test (n = 10). BP exposure, which induced approximately 30% heart rate reduction, did not affect mRNA expression of genes related to cardiomyocyte development: hand2 (D) and nkx20.5 (E). Only BPF and BPAF significantly downregulated bmp4 mRNA expression (F), based on the one- way ANOVA test followed by Dunnett’s post hoc analysis (n = 3). SC: DMSO 0.05%, BPA: 25 µM, BPF: 60 µM, BPAF: 4 µM, BPB: 12 µM, Mix 1: BPA 6.6 µM + BPF 13.6 µM + BPAF 1 µM + BPB 3.2 µM. ns, P ≥0.03; *, P ≤0.03; **, P ≤0.002; *** , P ≤0.0002; *** *, P ≤0.0001. The data are presented as means ± standard deviations.

Article Snippet: Embryos were stained with mCherry Mouse Polyclonal Antibody (Cusabio, Houston, TX, USA), Goat Anti-Mouse IgG (H+L) Highly Cross-Adsorbed Secondary Antibody, and Alexa Fluor Plus 594 (Invitrogen) according to a previously described protocol (Hammond-Weinberger and ZeRuth, 2020).

Techniques: Confocal Microscopy, Expressing

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: A structural peculiarity of Antarctic fish IgM drives the generation of an engineered mAb by CRISPR/Cas9

doi: 10.3389/fbioe.2024.1315633

Figure Lengend Snippet: Generation of the engineered 9E10 hybridoma cell line. Flow cytometry dot plots show, after electroporation without (A) or with pX458, containing the gRNA2, and the donor construct (B) , cell viability and mCherry expression (C,D) . Negative controls (A,C) (CTR-) are reported. Data from four independent experiments are presented as mean percentage of mCherry positive cells relative to the negative control (CTR-) (E) . *** p < 0.001 (two-tailed Student’s t test).

Article Snippet: For mCherry detection, the membrane was incubated with a 1:1,000 mouse anti-mCherry monoclonal antibody (Elabscience E-AB-20087) overnight at 4°C, followed by a 1:15,000 diluition of sheep anti-mouse IgG HRP-conjugated secondary antibody (Bethyl, A90-146P) for 1 h at RT.

Techniques: Flow Cytometry, Electroporation, Construct, Expressing, Negative Control, Two Tailed Test

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: A structural peculiarity of Antarctic fish IgM drives the generation of an engineered mAb by CRISPR/Cas9

doi: 10.3389/fbioe.2024.1315633

Figure Lengend Snippet: Schematic representation of engineering of the murine IgG1 heavy-chain gene locus. The region of the wild-type (WT) mouse IgG1 heavy-chain constant gene locus involved in genome editing. The gRNA targeting the intron between the CH1 and hinge exon (H) is indicated (A) . Engineered IgH locus resulting from the HDR-based exchange of the H sequence of mouse IgG1 with that of Antarctic fish IgM (B) . The coding sequence of the insert contains the Antarctic fish hinge exon in yellow, followed by mouse IgG1 CH2 and CH3 exons, linked through a linker sequence (L) to mCherry, used as a selection marker. The stop codon is reported in white. The donor construct is flanked by the homology arms (5′HA and 3′HA, green lines) of 1,005 and 2,421 bp, respectively.

Article Snippet: For mCherry detection, the membrane was incubated with a 1:1,000 mouse anti-mCherry monoclonal antibody (Elabscience E-AB-20087) overnight at 4°C, followed by a 1:15,000 diluition of sheep anti-mouse IgG HRP-conjugated secondary antibody (Bethyl, A90-146P) for 1 h at RT.

Techniques: Sequencing, Selection, Marker, Construct

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: A structural peculiarity of Antarctic fish IgM drives the generation of an engineered mAb by CRISPR/Cas9

doi: 10.3389/fbioe.2024.1315633

Figure Lengend Snippet: Validation of the production of anta-mAbs. Coomassie Brilliant Blue-stained 10% SDS-PAGE run under reducing conditions of purified WT and anta-mAbs. The L chain, the engineered and WT H chain bands are labeled on the right (A) . Western blot analysis using a mouse anti-mCherry monoclonal antibody as the primary antibody, followed by a sheep anti-mouse IgG HRP-conjugated secondary antibody (B) . The expected band at 75 kDa, corresponding to the engineered H chain, carrying the mCherry protein in anta-mAb, was detected. Molecular weight markers are shown on the left-hand side of each panel.

Article Snippet: For mCherry detection, the membrane was incubated with a 1:1,000 mouse anti-mCherry monoclonal antibody (Elabscience E-AB-20087) overnight at 4°C, followed by a 1:15,000 diluition of sheep anti-mouse IgG HRP-conjugated secondary antibody (Bethyl, A90-146P) for 1 h at RT.

Techniques: Biomarker Discovery, Staining, SDS Page, Purification, Labeling, Western Blot, Molecular Weight